Fukuoka H, Ogawa T, Minami H, Yano H, Ohkawa Y
Chugoku National Agricultural Experiment Station, Hiroshima, Japan.
Plant Physiol. 1996 May;111(1):39-47. doi: 10.1104/pp.111.1.39.
cDNA clones for two isogenes of nitrate reductase (NR) have been isolated from rapeseed (Brassica napus L.) androgenetic haploid embryos induced by microspore culture. NR mRNA accumulation can be detected by northern hybridization at 14 d after culture initiation when embryos develop to the heart/torpedo-shaped stage. Whole-mount in situ hybridization experiments demonstrate that the mRNA accumulation is developmental stage specific. In addition, even when cultured in media containing no nitrate, embryos accumulated NR mRNA to almost the same level as the control. This indicates the unique regulation of NR in embryogenesis in which NR mRNA transcription is activated in a developmental stage-specific manner that is independent of nitrate induction. In zygotic embryogenesis, a stage-specific accumulation of NR mRNA was also observed. By contrast, the obvious effect of nitrate on NR expression that has been reported in many plant species was also confirmed in rapeseed leaf. Quantitative combined reverse transcription-polymerase chain reaction analysis suggests that the flexible and variable regulation of NR expression, which is organ specific, nitrogen metabolite specific, and developmental stage specific, is caused principally by regulation of one major structural gene.
已从通过小孢子培养诱导的油菜（甘蓝型油菜）雄核发育单倍体胚胎中分离出硝酸还原酶（NR）两个同基因的cDNA克隆。培养开始14天后，当胚胎发育到心形/鱼雷形阶段时，通过Northern杂交可检测到NR mRNA的积累。整体原位杂交实验表明，mRNA积累具有发育阶段特异性。此外，即使在不含硝酸盐的培养基中培养，胚胎积累的NR mRNA水平也几乎与对照相同。这表明NR在胚胎发生过程中具有独特的调控机制，其中NR mRNA转录以发育阶段特异性方式被激活，且独立于硝酸盐诱导。在合子胚胎发生中，也观察到NR mRNA的阶段特异性积累。相比之下，许多植物物种中报道的硝酸盐对NR表达的明显影响在油菜叶片中也得到了证实。定量逆转录-聚合酶链反应分析表明，NR表达的灵活可变调控具有器官特异性、氮代谢物特异性和发育阶段特异性，这主要是由一个主要结构基因的调控引起的。
